The retroviral enzyme, integrase (IN), catalyzes the insertion of newly synthesized viral DNA into the host cell chromosome. This event is required for normal viral replication. Thus, like HIV-1 reverse transcriptase and protease, IN is a potential target for inhibition in prevention or treatment of AIDS. The major goal of research described in this proposal is to understand the molecular structure of HIV-1 IN, alone and in complex with its DNA substrates, so that design of inhibitors may build upon a rational foundation. Three Specific Aims are proposed: The first Specific Aim describes structural analyses of distinct domains and full length IN, in the presence of metal cofactors, selected model substrate inhibitors and monoclonal antibody fragments, using X- ray crystallography and NMR. The second Specific Aim will test specific models of IN organization and function suggested by the structural data, using site-directed mutagenesis and chemical modification of IN. The third Specific Aim describes the use of a novel, synapsed-end viral DNA substrate to investigate how IN interacts with DNA in a functional complex that catalyzes coordinated processing reactions. A specific model that predicts DNA bending and partial unwinding will be tested, using DNA footprinting techniques. Experiments described in this proposal employ a broad range of state-of-the-art molecular genetic, biochemical, and biophysical methods, as well as novel substrates and immunological reagents. They also build upon the extensive experience of the PI and collaborators with both the human (HIV) and avian (ASV) viral integration systems, and the ability to utilize ASV IN as a valuable base of comparison. The knowledge gained should help to delineate features that are specific to HIV. The results of these studies provide new molecular targets for inhibition of HIV integration and development of AIDS therapies.